CHAPTER ONE INTRODUCTION
1.1 Introduction
Leeching or bloodletting has been long introduced since the ancient time as one of method to cure several diseases believing that it could exonerate noxious substances produced by these diseases. However it soon diminished around 20th century in Europe as antibacterial therapy takes place and soon becomes an unpopular until now. The revived of leech therapy in modern medicine happens in 1970’s with limited application in the usage of relieving venous congestion in microsurgeries. Up until now scientist and medicine practitioner still making discoveries of this amazing ability of leech for many more functions especially in therapeutic areas.
In Malaysia, Leech therapy is considered as alternative medication that believed can cure or relieved some of the ailment that has been practiced since long time ago.
Therefore, the effect of body composition is one of the methods of assessment to evaluate whether the therapy that has been practiced since their ancestor time is a worthwhile tool for therapy in healing some said illness. Using impedance method for body composition measurement can determine the constituent of body changes which model as 2 important compartment that divides the body into fat mass and fat-free mass (FFM). This measurement is normally used by nutritionist as this invasive method is very suitable to determine the health of a person which directly affected by the composition of the fat in the body that can be further subdivide into water, mineral and protein constituents. The result could be used to measure the effectiveness of the leech therapy treatment as the readings and changes are taken into consideration before and after the treatment.
2 1.2. Research Problem and Problem Statement
1) The effectiveness of leech therapy used as medicinal treatment is not
quantitatively proven and thus need to be highlighted as the growing interest towards the ability of leeching is widely explored.
2) Leech therapy need to be measure quantitatively using an engineering tool as method of assessment of the effectiveness of the therapy.
1.3 Objectives
1) Measure and represent the data of the body composition of patient that undergo leech therapy treatment
2) Analyse the result of body composition of the leech therapy patient using statistical tool
1.4 Hypothesis
1) Leech therapy treatment will have some changes on body composition 2) Effectiveness of leech therapy treatment can be represented quantitatively.
1.5 Scope of the Study
Scope of this study is restricted to Malaysian patient that undergo the leech therapy treatment and only immediate response of the treatment is measured.
1.6 Significance of the Study
Mere believe in the alternative medicine needs to be proven quantitatively so that people can be convinced and not deceived by unsupported claims. As leech therapy exist since long time ago, accreditation is necessary to assess the effectiveness of this method.
CHAPTER TWO LITERATURE REVIEW
2.0 Modern Medicine vs. Complementary and Alternative Medicine (CAM) Modern medicine is view as a mainstream and conventional ways based on scientific evidence while alternative medicine has been define in many terms like holistic, traditional and mostly categorized as complementary and alternative medicine (CAM). Treatment of diseases has been developed and modernized with a lot of new tools, surgery and pills for either preventing or curing purposes. However, alternative medicines is still being practice and recently have gain a lot of attention from all over the world. It can be divided into diets and supplements, botanicals products, unconventional agents, traditional medicine and energy healing (Cassieth et al., 2001).
If to compare both of these methods, modern medicine is still considered the main method used for healing as the governments also in support of this kind of treatment as it is based on scientific value and proven to save many life every day. However, CAM is a minority but gaining its popularity day by day and it could be revive and used as conventional medicine if scientific research can support the effectiveness of these methods. Therefore, research such as changes in body composition to check the effectiveness of the traditional medicine like leeching can contribute to the scientific evidence CAM as a whole so that it could be accepted by the world community.
4 2.1. Modern medicine
2.1.1 Surgery
Surgery has been a mean of doctors in getting rid of or fixing out affected area that has been diagnosed ‘ill’. From cosmetic surgery, teeth and transplantation or removing any tumours has been common nowadays. This procedure has saved many lives especially when it involves a vital organ like heart that has many possibilities for getting weak or failed like due to multivessel disease suffered from the diabetic patients (King, 2005). Bypass surgery or known as coronary artery bypass graft is normally performed to reduce the risk of death from coronary artery diseases. At the same time, injections and surgeries are inseparable combo a surgeon used as a tool in the operation.
Injections not only used as pain reliever it could also be used to inject certain serum or protection agent to human body.
2.1.2 Pill/ tablets/ capsules
Pills, tablets or capsules that contain drugs to prevent or cure diseases are most necessary thing that a doctor will prescribed to its patient. Thus, a lot of research on drugs that can act on certain functions of our body has been actively pursued to treat various different of illness. The modern practitioner believed that drug must be assesses in the patient level. A report by WHO stated that drug utilization needs to have answer to some assessment questions like why, who the prescriber and person been prescribed also whether the patient medicine intake is in order and what are benefits and risk of the drugs being used (WHO, 2003). In addition, these are all important since a strict qualification by the food and drug administration always ensure that the drug used comply with some basic rule for it to be used in public.
2.1.3 Medicine tools
Modern medicine is also in another level with the engineered equipment and tools to assist the physician/ therapist and doctors in handling and diagnosing diseases.
Application of ultrasound, nuclear magnetic resonance or x-ray in imaging system brings a great advancement to the modern medicine as a whole. These tools continue to flourish along with the modern medicine as a lot of potential been put aside for the scope of imaging in medicine. Besides that, tools like lasers not just benefit the physicist but give advantage in medicine world with the new ways of making surgery with lasers. Three basic laser principles that have been used in developing tools for medicinal purpose are photochemical effects, absorption which converted into heat and non-linear effect that can remove material fragment without heating the surrounding tissue (Senz and Muller, 1989). These are already applied in recanalization of arterial blood vessel, fragmentation of kidneys, bladder, lasik surgery and many more.
2.2 CAM
2.2.1 Diets and supplements
Some of the dangerous illness like cancer can actually be cure not just by the means of conventional medicine. Therefore, there are some theories in healing the cancer by using diet and eating supplements that is considered a natural way to fight cancer. This is because some foods contain a lot of nutrients that can boost the immune system. By eating the correct foods with a strict diet a person is believed to be able to fight the non-rapid cancer tissue by slowing the progression of cancer and strengthen the patient’s body (Miller et al., 2000). Moerman diet therapy is popular in Netherlands with 2 important parts where the main diet involves a lot of organic product and also the supplement foods. There are eight key nutrients in this diets namely Vitamin A, B complex, C, E, citric acid, iodine, iron and sulphur (Kehr, 2011). This non-toxic method
6 receives a lot of attention as it can also be applied to healthy person to keep fit and prevent cancer in a better way. Other types of diets available are Houtsmuller diet, Macrobiotic diet, Gerson diet and a lot more.
2.2.2 Botanicals products
Besides diet and supplements, botanical products like herbs have been practice since long time before especially Chinese herbal medicine products. Sometimes, old generation will take the herbal medicine as a daily dose as raw or processed herbs with variety mixture of ingredients that believed can bring good health and youth to them.
Malay herbs like kacip Fatimah, tongkat ali and hempedu bumi has even revolutionized as a commercial product so that it can be find easily (Aziz et al., 2005). Chinese traditional herbs also has been popular not in Asia but in western country and a lot of research on natural products from these herbs is made for cancer therapy treatment.
Molecular target for the treatment are mostly signal transduction protein like kinases which able to control the malignant processes in cancer cells (Efferth et al., 2005).
2.2.3 Unconventional agents
Some of the alternative medicine falls under the category of unconventional agents such as shark cartilage, Ozone therapy, Hasumi vaccine, Di Bella therapy and more of new method that is unconventional. Shark cartilage has been an interest since research of glycoproteins that can extend the life in leukemic mice where extracted from the hammerhead sharks (Petit G.R. and Ode R.H., 1977). Moreover, there is also study done that shows that intraocular implantation of shark cartilage pallets can repressed tumour (Gawler I., 1984). Besides that, ozone therapy is more popular in Malaysia where the treatment could be used to cure external wound and even in dentistry. Since ozone has various effects such as antimicrobial, analgesic, immune stimulating and a lot
more benefits, this treatment has a variety of way to be used as cure (Seidler et al., 2008). However, some precautious actions need to be taken so that any complications should be avoided.
2.2.4 Traditional medicine
Traditional medicine is one of the famous methods in CAM as it has been practiced since ancient time and still has a demand from one generation to another.
Some of the famous traditional medicines used are Acupuncture, Hirudotherapy, Ayurveda, Qi gong and others. Acupuncture works through the theory of healing the
“energy” called “Qi” inside the body via a network of pathways called “Meridian”. The 365 acupuncture points are assessed through the 14 primary meridians which correspond to 12 different organ systems in our body. The mechanism is vague but some explaining that placing needles in the acupuncture points will stimulates a signal in the connective tissue. This signal will activate a biomechanical or biochemical reaction that helps in healing of body (Povolny, 2008). Besides that, hirudotherapy or leeching is a famous method in bloodletting and will be elaborate more in the next topic.
Hirudotherapy is one of the ways in balancing back the biological humour according to the Ayurveda treatment in which there are three humours in the Indian traditional medicine referred as vata, pitta and kapha.
2.2.5 Energy healing
The energy healing is another types of alternative medicine that could cure illness. Some of the examples are homeopathy, bioresonance therapy, teletherapy, urine therapy a much more methods. Homeopathy for example has been introduced since 200 years ago by a German physician by using the “Principle of Similar”. The principle works by giving out medicine with matching drug symptom to the symptom in the
8 patient in the minimum dosage (Jonas et al., 2003). Bioresonance therapy is another types of therapy that has been invented in Germany using an electromagnetic concept.
This therapy is believed to be able to stimulate a change of bioresonance in cell so that the cells could be healthy again after affected by changes carried by the disease.
Therefore, the good energy is once restored and this claim is used for other types of energy healing agent too.
2.3 Leeching/ hirudotherapy in ancient time
Leeching has been an old remedy since 1500 BC where it is found in the painting in an Egyptian painting displaying the usage of leech as medicine. During the Roman Era, leech has been promoted because they believed that bloodletting will clear their bodies from the harmful substances produce by diseases. This faith of them is performed so that they can restore the four humour of human body namely blood, phlegm, yellow bile and black bile for proper balance (Munshi et al., 2008). Then, it becomes extremely popular during the 18th and 19th century where indication of leeching is used in acute laryngitis, nephritis, subacute ovaritis, epitaxis, ophathalmia and brain congestion (Adams, 1988). Some prefer using leech than cupping as they can be used in the delicate areas like eyes, gums, breast and testicles. Later it becomes less popular as new modern medicine emerge until late 19th and early 20th century where some scientific research has found some usefulness of leech. For example, Haycraft in 1880 discovered the antithrombotic properties of leech saliva and Jacoby in 1904 has found the potential anticoagulant factor named Hirudin (Fields, 1991). This is expanded to the usage of leech as hirudotherapy in plastic, reconstructive and trauma surgery.
Furthermore, nowadays leech is still on research to be used in many medicinal applications as it has high potential in helping to cure variety of diseases.
2.3.1 Leeches
For about 650 species of leeches, only some are used in the therapies like Hirudinaria medicinalis, Hirudinaria asiatica and Hirudinaria manellensis. Leech normally can be found on the fresh water and feed on decaying plant material. Leech is one of the animals under the category of hermaphroditic which has 2 reproduction organs both male and female. However, leech requires another leech to reproduce and lay egg in cocoons (Mory et al., 2000).
The leech can be categorized as annelids which looks like rings or segmented body same as the ringworms or earthworms. The medicinal leech or scientifically called as Hirudo medicinalis is quite special in which it can cause the deepest bite and eruption of blood after treatment for a longer time than others (Irish et al., 2000). Being a Hirudo medicinalis, this type of leech can expand themselves three times more than its resting length. There are about five segments for each of the 102 annuli in this leech which has a posterior sucker triradiate with three jaws to bite and smaller anterior sucker for feeding (Valauri, 1991).
Figure 2.1: Leech
(Source: http://www.digitalmediatree.com/library/image/88/leech.jpg) [Viewed : 12 September 2011]
10 Based on the picture, leech will use the jaw to bite and these jaws are connected to the pharynx, esophagus and crop. Crop is where it is literally said as the leech stomach that can be expanded during the feeding. The blood is store inside the crop and can be maintain in fluid state inside their bodies. The crop is connected to gizzard that finally connected to the intestenum and ends at the posterior sucker. The medicinal leech is used for therapy as the therapeutic effect can be obtained from the continued bleeding after the leech is removed. The amount of blood from the bloodletting process is approximately 5-12 ml and the bleeding from patient shows a good sign where this happen due to anticoagulant properties from the leech salivary gland secretion (Engemann and Hegner, 1981). Some animals are actively synthesizing the salivary anticoagulant to secure their nutritional requirements of the body which includes leeches (Ribeiro and Garcia, 1981). Ribeiro, 1981 also state that the salivary active compound could prevent blood finding or blood feeding in some animals.
The way the leech feeds is mostly stimulated by mammalian range temperature and the content of sodium and arginine in blood (Lent, 1986). In general leeches do not just feed on human blood. Some are actually able to consume small invertebrates and some will suck blood using their strong attaching sucker to the host. They have some preferential in blood from certain species and considered particular in feeding pattern.
Since not all leeches have jaws, some can secrete enzymes that help them attach to an opening through the skin. In most of the cases, the host would not be able to realize the leech attack until the end because the leech can produce natural anesthetic substance emanate from the saliva of leech. The host will have bleeding for a period of time after the process since the saliva also contains anticoagulants known as hirudin that catalysed conversion of fibrinogen to fibrin and thus prevent blood clotting (Mory et al., 2000).
Blood remains in fluid state as the crop of leech is colonized by an endosymbiotic
bacteria like Aeromonas spp to help leech digest the blood that they feed (Eroglu et al., 2001). Table 2.1 summarized the major enzymes that are important in leech:
Table 2.1: Major enzymes important in leech
Enzyme Function
Hirudin Act as anticoagulant
Bdellin Protease inhibitor thus act as anti- inflammatory
Apyrase Powerful platelet anti-aggregate to make blood flow more fluid
Eglin Inflammation inhibitor and antioxidants
Destabilase Powerful platelet anti-aggregate to dissolve blood clot Hyaluronidase For diffusion and antibiotic
Lipase and Esterases For hyperlipidemia
Anti-Elastase Degrade cutainous elastin particularly at the level of skin Vasodilatory Similar to histamine (act as neurotransmitter)
Catacholamines Act on the nerve terminator (specifically at skin)
2.3.2 Mechanism of Hirudotherapy
The mode of action of hirudotherapy depends more on the secretion of enzymes from the salivary glands of leech. The salivary glands produce a lot of active substance that act as anticoagulant, anti-inflammatory, bacteriostatic and analgesic action.
Therefore, the effect of these could eliminate hypoxia, increase immunity, restore any damaged vascular and reduce the blood pressure. In addition, the medicinal leech can directly act upon cellular and plasma factors associated with blood clotting as it blocks the attachment of thrombocytes and conversion of fibrinogen to fibrin.
12 2.3.2.1: Application in Soft Tissue Hematomas
Hematoma can be called as the phenomenon where collection of blood localized outside the blood vessels. This could happen in any part of body including the oral system in which macroglossia could happen due to seizure- related activity. The extensive lingual swelling could also come from penetrating injuries from the lower face or bleeding from the fracture site (Grossman and Karlovitz, 1998). To prevent acute airway emergency, treatment should be done earlier to avoid worsening swelling due to congestion in venous and lymphatic system. Thus, hirudotherapy has been reported as one of successful method in handling post-traumatic lingual swelling.
Heckmann et al., reported that there is significant improvement over the subsequent 24 hours after the treatment of using 13 leeches sucking out about 145 ml blood (Heckmann et al., 2005).
2.3.2.2: Application in Tissue Flap Reconstructions
Tissue flap reconstruction is one of the most popular applications of hirudotherapy as it is effective in removing stagnant blood from a skin flap during reattachment of limbs. The stagnant blood is necessary to be put out of the area affected as pooling of blood will lead to the increases of venous pressure. If this is prolonging, the wound will not be provided with oxygen and nutrients thus the injury will not heal.
So, in this case hirudotherapy come in handy because the leeches can absorb all the stagnant blood and after 3-7 days the damaged veins will resume the normal colour and pressure so that blood can flow to the affected area continuing the wound healing process (Amrit, 2010). Figure 2.2 depicts the process of application, attachment and satiation of leech in a facial flap reconstruction.
Figure 2.2: Application of hirudotherapy to tissue flap reconstruction (Source: Porshinsky et al., 2011)
2.3.2.3: Application in the Setting of Severe Soft Tissue Injury and Surgical Replantation
The venous disorder study has been made from 1995 to evaluate effectiveness using hirudotherapy for patients that have difficulties due to venous obstruction after head and neck tissue transfer (Chepeha D.B., 2002). All the flaps from the 8 patients could not be saved using conventional surgical or thrombolytic therapy. Therefore, after 6.6 days for inosculation and duration in intensive care unit for 9.6 days, it bring successful result where the flap is save by using about 215 leeches per patient.
In most of the reconstruction cases, the severe soft tissue injury are due to venous congestion where this is detect from a few symptom like development of tissue edema, purple discoloration and warmth of the reconstructed or replanted tissue (Hullet et al., 2007). Applying leech to the affected area will reduce the swelling and discoloration and thus venous congestion will decrease at the same time.
Yet, another procedure that has been using hirudotherapy which is suitable for sensitive area is penile amputation. This has been a good alternative since penile replantation requires adequate postoperative venous outflow (Mineo et al., 2004).
During the surgical procedure careful approximation of urethra and corporal bodies of the dorsal vein plays an important role. Therefore, for non-microsurgical replantation procedure, application of leech is one of the factors for the success of the replantation
14 (Mineo et al., 2004). Some of the common effects from this non microsurgery procedure are skin loss, urethral strictures and fistulae, loss of glans penis also sensory deficits (Bhanganada et al., 1983). So, hirudotherapy play important steps that take place after the operation. In figure 3 the replantation is a success after using the leech where from a to b the leeches are held until the attachment and c shows that leech already attach and start to increase in size until it satiate from d to e.
Figure 2.3: Application of hirudotherapy in a case of digital replantation (Source: Porshinsky et al., 2011)
2.4 Body composition
A body composition is the measures to describe compartments of our body that consists of fat and non-fat component (water, protein, minerals). This measurement is important as it can gives information on the components of human body due to nutrition, diet, exercise, disease and even therapy activities. Some of the basic definitions that must be known in body composition are:
1) Body density (Db) : total body mass expressed relative to total body volume 2) Body mass (BM) : Measure of the size of the body
3) Fat mass (FM) : All extractable lipids from adipose and other tissue in the body 4) Fat free mass (FFM): all residual lipid-free chemicals and tissues including
water, muscle, bone, connective tissue and internal organs.
5) Lean body mass (LBM) : Fat free mass plus essential lipids
6) Relative body fat (%BF) : Fat mass expressed as a percentage of total body weight
7) Total body bone mineral (TBBM) : A measure of the osseous mineral content of the body
8) Total body mineral (TBM) : A measure of the osseous and non-osseous mineral content of the body
9) Total body water (TBW) : A measure of the intracellular and extracellular fluid compartments of the body
In order to measure the body composition of human body, measurement should be invasive and convenient where estimation can be done indirectly. Thus, early studies have been developed to model the body composition that divide the weight into two or more components. There are several applications of body composition:
1) To identify health risk due to extremely high or low level of total body fat 2) To monitor diseases by changes in body composition
16 3) To assess effect of nutritional intake, dietary, therapy and exercises to body
composition
4) To keep track of healthy body weight for athletes
5) To monitor children development from changes in body composition
Therefore, body composition is one of the methods that is convenient and used frequently by many practitioner to see changes in body composition.
2.4.1 Body composition models
There are a few models that have been proposed since early studies in which they are:
1) Two component model (2-C) 2) Three component model (3-C) 3) Four component model(4-C) 4) Six component model(6-C)
Out of these four models, the 2-C model has been generally used. However, by using isotope dilution, dual energy x-ray and neutron activation analysis it is possible to derive the other models which can quantify and accurately measures body composition that divide fat free mass (FFM) into water, mineral and protein (Boileau, 1977).
2.4.1.1 Two component model
This model has been developed by Brozek and Siri where the model divides the compartment of human body into FM and FFM. They find and formulate a relationship to calculate the %BF. However, some assumptions are taken into account where:
1) Densities of FM and FFM are additive as well as constant for all people.
2) The content of FFM proportion is constant within and between all people.
3) Each different people only vary in term of fat and obesity tissue.
2.4.1.2 Three component model
This is enhanced from the two component model where body weight is the summation of fat, water and solids which consists of mineral and protein. The model developed is considering the hydration factor that is different from one individual to the other. However, Lohman also proposing the three components model where body weight is made of fat, mineral and protein with water. These two models are representing the body composition on molecular level while there’s another three component model which is on tissue/cellular level. This is done by using the dual- energy X-ray absorptiometry that divides the body weight equation equal to summation of bone, bone free lean tissue and FM. The model which develops by Ellis separates the equation into two 2-C model. One is for bone and soft tissue mass and the other is lean and fat tissue.
2.4.1.3 Four component model
The four component model is based on the composition of body weight is made of fat, water, bone, mineral and protein. This has been developed by several researches like Friedl, Selinger, Heymsfield and Baumgartner. This model can portray estimation of fat more accurate than 2-C model but requires more measurement on different variable.
2.4.1.4 Six component model
The six component model is more detailed than the other types of model as the body composition is assess in atomic level. According to this model, body weight consists of TBW, TBN (total body nitrogen), TBCa (total body calcium), TBK (total body potassium), TBNa (total body sodium) and TBCl (total body chloride). This kind
18 of measurement need direct analysis of chemical composition in vivo. The atomic major elements of body are measured using neutron activation analysis.
2.4.2 Body composition measurement
There are a few types of assessment where we measures body composition by using the method of Hydrodensitometry, Anthropometry, Bioelectrical impedance analysis, Air displacement Plethysmography, hydrometry, Dual energy X-ray absorptiometry (DXA), Neutron activation analysis, Whole body counting of potassium, Computed tomography, and magnetic resonance imaging. These methods are developed to identify and estimate the body composition in molecular or atomic level.
2.4.2.1 Hydrodensitometry
Hydrodensitometry is also known as hydrostatic weighing or underwater weighing in which it is used to measure the body volume. It works by the principle of Archimedes where the estimation of total body volume is equal to the water displaced by the body when it is fully submerged. However, it is not practical to collect the amount of displace water thus measurement is done by determining the underwater weight (UWW) of an individual. This is normally difficult to conduct for excessively obese, pregnant, elderly or disabled subjects (Brodie and Slade, 1990). This method requires a sensitive and continuous measurement through the level of exhalation that will give the value of readings. The readings are the estimation of UWW which is directly proportional to the volume and weight of the water displaced by body volume.
2.4.2.2. Anthropometry
This method has been used to predict the body density that can be change to percentage of body fat. It involves skinfold technique where the thickness of skin fold is measure at different part of body, including bone dimension and limb circumferences.
This technique is very common in the lab and anthropometric field for assessing the body fat in body composition and nutritional evaluation (Sloan, 1967). The skinfold method is use as it can estimates the general fatness and the distribution of subcutaneous adipose tissue which assume to represent the total body fat. Normally, the potential sources of error come from the caliper selection and tester reliability in which it fails to estimate simultaneously all possible sources of measurement error (Brodie, 1998). The error can also result in different degree of error variance for different conditions (Sloan and Shapiro, 1972; Lohman et al., 1984).
2.4.2.3 Bioelectrical Impedance
Bioelectrical impedance analysis (BIA) method is used since 1980’s and now famous for practitioner to make different diagnosis and make a long term approach to treatment. Using the tools for measurement, BIA will give the result in impedance value of body tissues. The impedance then can be calculated to estimate the total body water (TBW) which also estimate the FFM and adiposity. In addition, BIA can also be used in approximating the body cell mass and TBW in different kind of clinical condition (Kyle et al., 2004).
20 2.4.2.4 Air displacement Plethysmography (ADP)
This method is an alternative to hydrodensitometry as it much more convenient and requires only minimal technician skill to conduct. The ADP is using a Bod Pod system where the derivation of BV and Db is based on the inverse relationship between
pressure and volume (Boyle’s law):
1 2 2 1
V V P P =
Figure. 2.4: Two chamber bod pod system (Source: http://www.iub.edu/~k536/measure.html)
[Viewed: October 14, 2011]
Figure 4 show the Bod Pod system which consists of front chamber and rear chamber that separated by a wall made of molded fiberglass. An oscillating diaphragm is attach to create changes in volume where the measurement is taken twice and pressure and volume is calculated before and after the patient is inside.
2.4.2.5 Hydrometry
Hydrometry is the measurement of body water that is the most abundant components in the body. This technique is based on the dilution techniques where a tracer will be diluted in one of the water components. Hydrometry requires two fluid samples which is normally blood, saliva or urine. The first sample is for baseline measure where the second one measures the concentration of the tracer. Then, the TBW is calculated from the estimation of the water components that already equilibrate with the tracer like tritium oxide, deuterium oxide and oxygen-18 (Heyward and Stolarczyk, 1996).
2.4.2.6 Dual energy X-ray absorptiometry (DXA)
DXA provide a measurement of total body bone mineral and bone mineral density. In addition, it also able to measures FFM, %BF and soft tissue mass where it is equal to the summation of lean tissue mass (LTM) and FM. The bone mineral content that is captured by the DXA is given in grams and projected area of the measured site is in centimeter (Adams, 2003). Then bone mineral density is calculated by dividing the bone mineral content by the bone area (Blake et al., 1999). The principle is based on the measurement of attenuation of x-ray with high and low photon energies. Since the X- ray is absorbed by the body, the attenuation of the energy varies depending on the densities and chemical composition of the tissue.
2.4.2.7 Neutron activation analysis (NAA)
This method can give details composition of body in atomic level where suitable for 6-C model. During the assessment, a beam of neutrons will be projected to the patient thus creating an isotope that emits gamma rays. Then components like calcium,
22 sodium, chlorine, phosphorous, nitrogen, hydrogen, oxygen and carbon can be determined through the measurement of the emission of the gamma rays.
2.4.2.8 Whole body counting of potassium
This method is able to give the value of body cell mass by the measurement of total body potassium. Potassium is an element that can be found in the intracellular with different types namely 39K, 40K and 41K. Only 40K can be detected by the gamma rays making use of it as counter to determine the whole lean mass of the body. Finally, the fat percentage in the body can be determined from the lean mass and total body weight.
2.4.2.9 Computed tomography
Computed tomography (CT) is also based on the difference of X-ray attenuation where it allows identification of bone, adipose tissue and lean tissue. It is a powerful radiographic tool where fatness can be measures by combining a specific part with the whole body. In addition, due to the properties of CT that is more sensitive to small changes in attenuation result in the ability of CT in producing image of soft tissue in great clarity compared to standard radiography (Brooks and DiChiro, 1975;
McCullough et al., 1976).
Figure 2.5: CT scan
(Source: http://www.iub.edu/~k536/measure.html) [Viewed: October 14, 2011]
The figure depicts the concept of CT scan where the x-ray tube is placed opposite poles of a large ring that rotate around the patient. Then the computer will generate the 3-D image of organs and tissues slices by slices using the lead shutter in front of the X-ray tube and X-ray detector after complete rotation of 360°.
2.4.2.10 Magnetic resonance imaging (MRI)
MRI similarly can produce an image and simultaneously detect subcutaneous and visceral fat. It gives an unbiased measurement of fat content, internal and subcutaneous for a large range of body shapes and sizes (Thomas et al., 1998). Using MRI, the patient will be placed in a magnetic field that aligns together hydrogen nuclei when radio pulsed frequency is applied. When the radio frequency is turned off, the hydrogen nuclei will emit the radio signal that they absorbed creating an image at tissue level.
2.5 Bioelectrical impedance analysis (BIA)
BIA is method that uses an alternating current (AC) to measure the opposition of body tissue. Normally, the impedance value that captured will be varying according to the frequency of the current used. There are a few ways of using BIA where sometimes, single frequency or multi frequency is used in the analysis.
2.5.1 BIA principles
BIA works on the principles that relate the conductor volume, length, resistivity and impedance value. The conductor principle can be compare to the human body where conductor itself is the human body which has volume (V), height (L), specific resistivity for components like FM and FFM (ρ), and body impedance (Z).
24 2.5.1.1 Body impedance (Z)
Impedance value is a resistive property of alternating current (AC) that is a frequency dependant. It consists of two components which are resistive (R) and reactive capacitive (Xc). The relationship can be put into an equation of:
2 2
2 X R
Z = c +
The value of Xc refers to the value of body capacitance that present in the body when AC is in contact with the cell membrane. Therefore, Xc can be related to the body cell mass (BCM) that contains active components of the body like muscle, blood and immune cells. Resistive value, R, is the indicator of the extracellular mass and intracellular mass in which R is assume proportional to FM or FFM. The value of R is a resistive value when a current flow through the body and can indicate that high reading of R shows that FFM content is low in human body and vice versa. The effect of Xc sometimes is negligible due to comparatively small value but becomes significant as the frequency increases.
2.5.1.2 Volume and body composition
The volume of a conductor can be interpreted by the equation: V = L×A. Since Length can be related to the height of human then area is derived from the relationship of resistance for a conductor:
A R ρL
= . Since R is proportional to impedance,
rearranging the equation result in Z A ρL
= . Thus, deriving again the equation of volume will result in:
Z V L
ρ 2
=
--- (1)
This volume can be interpreted as the volume of the FFM or TBW that directly proportional to square of height and inversely proportional to value of impedance of body. However, this equation holds an assumption that the conductor is a perfect cylinder with a uniform cross sectional area. This is not true as a whole as human body can be represented as five cylinders excluding head (Kushner, 1992). All the five cylinders are different in cross sectional area resulting in different measurement of resistance (Baumgartner et al., 1989). Thus, to cater all the differences in the body
segment, the equation of volume become:
trunk arm
leg z
L z
L z
V L
+
+
=
2 2
2
2
2 ρ ρ ρ
.
2.5.1.3 Calculation of body composition
There are a few formulas in calculating the body composition components like TBW, FFM, FM, and % BF. Using BIA method, these readings can be measured at the frequency of 50 kHz using the formulas below (Lukaski, Bolonchuk, 1988):
(
sex) (
weight) (
age)
ce sis height
TBW 3.05 0142 0.069
tan 372 Re
. 0
2
− +
+
=
--- (2)
• Height is in cm
• Sex: Male = 1, Female = 0
• Weight is in kg
• Age is in years
73 . 0 FFM =TBW
--- (3)
• 0.73 is known as the hydration constant FFM
Weight
FM = − --- (4)
% 100
% = ×
weight BF FM
--- (5)
26 2.5.2 BIA models and approaches
There are a few models and approaches that have been used in determining the TBW and FFM in the body for BIA. However, the concept still applies the same where two electrodes namely source and sink is used to determine the current flow pass through it. The current that pass through the body will go through all the conducting material and voltage drop across the source and sink can be measured. These conductors are normally in the form of charged ions like sodium and chloride ions.
2.5.2.1 Single frequency BIA (SF-BIA)
In this method, only one frequency which is at 50 kHz is used to measure the impedance across the electrode that is paced on the hand and foot. However, some are placing the electrodes across hand to hand and even foot to foot (Jebb et al., 2000; Utter et al., 1999).
2.5.2.1.1 Series model
This is one of the traditional models where the assumption is considering only one conducting path and human body consists of series of resistors. Using a single frequency of 50 kHz, the impedance value will be used to determine the TBW and FFM.
However, this method is only suitable for patient with normal ratio of ECW to ICW.
2.5.2.1.2 Parallel model
This model made of assumptions where the body is not just a series of resistors but parallel as it suits more of the human physiology. The parallel model is the reciprocal of the series model and preferred to be used for patient with fluid imbalance as it is suitable for estimation of ICW and BCM.
2.5.2.2 Multi-frequency BIA (MF-BIA)
Since single frequency BIA cannot penetrate cell, multi-frequency BIA can achieved more in order to estimate the FM, ECW, ICW and TBW. The range of frequencies start from 0 up to 500 kHz is used to give more accurate and less biased value than SF-BIA.
2.5.2.2.1 Bioelectrical impedance spectroscopy (BIS)
The BIS is more to prediction using the mathematical modelling and mixture equation. The mixture equation like Cole model can show improvement in accuracy than regression approach as BIS utilizing correlation between R and body fluid compartments. From there, prediction equation can be derived provided reliable data fitting and valid fluid distribution model is included (Kyle et al., 2004).
2.5.2.2.2 Segmental model
The value of impedance is greatly influenced by the cross sectional area of the body.
Comparing the five model cylinder, trunk will represent most of the FFM value than others. Therefore, segmented model can give more accurate value than the whole body BIA. Segmental BIA is able to calculate the distribution of body composition segment by segment thus useful in patient with distorted fluid distribution. In addition, to used segmental model requires additional two more electrodes in any part like torso, leg or arm.
28 2.5.2.2.3 Localized bioelectrical impedance analysis
Localized BIA is also one of the methods that can give specific segment value.
As the traditional BIA concluded population specific, localized BIA can be more focus to avoid the interference effect for example in measuring the local abdominal fat by using the localized BIA (Scharfetter et al., 2001).
2.5.2.2.4 Bioelectrical impedance vector analysis (BIVA)
BIVA is type of analysis that is consider as a standalone model that can give direct assessment of patient from direct impedance measurements plotted in bivariate vector standardized by the subject’s height. This method is useful and advantageous in the clinical settings as the result is not biased due to choice of regression equation since no conversion of raw data to body composition is required. The result can be plot in R- Xc plane and stratified accordingly due to different vector distribution pattern (Piccoli et al., 1995; Piccoli et al., 2002).
2.6 Statistical test
Statistical test is important in organizing data and making conclusion from the raw data. It’s a powerful tool in which one can represent a set of raw data in graphical or tabulated form and relate the variable while making estimation and get better understanding of situation of the data portrayed. Some of the statistical analysis that is common to researchers is t-test and analysis of variance (ANOVA) that involves the description of mean.
2.6.1 T-test
T-test is a test which able to determine the separateness of two sets of measurement. Therefore, mean or average value and distribution can be depicted by using t-test. There are three types of t-test which is independent t-test and dependent t- test and one sample t-test.
2.6.1.1 Independent t-test
This test is applicable for comparing two different population or sample for one variable. Normally, for a simple experiment the result of the experiment and the control group can be compared and analyse by using the independent t-test.
2.6.1.2 Dependent t-test
The dependent t-test is used to evaluate data by comparing average of one single group for two different variables. This is normally called as paired sample test as the result from the two mean is from the same group of interest. The dependent t-test usually been applied in before and after situation where the effect of the variable is determine by comparing the mean.
30 2.6.1.3 One sample t-test
This type of test is more direct as we only compare mean of a variable with a constant value that is usually a hypothesized value that already calculated before the test is conduct. It can determine the validity of the measurement or maybe directing to another conclusion of the test.
2.6.2 ANOVA
ANOVA is more powerful than t-test as it can compare means more than two groups. The reliability of t-test is valid up until two groups thus urging the ANOVA method to be used to get result and see the relationship from different group or variables.
Generally, there are two main types of ANOVA namely one-way ANOVA and two-way ANOVA.
2.6.2.1 One way ANOVA
One way ANOVA is the upgraded version of independent t-test where it can evaluate a mean for more than two groups. Sometimes, in real situation cross evaluation for several samples is required to get a confirmation of the hypothesis of an experiment.
2.6.2.2 Two-way ANOVA
This method is different in terms of evaluation of mean can be compared with two different variable for the same group. This will allow the researcher to determine the combined effect of the independent variables plus assessing the interaction between the variables in influencing the mean on the dependent variable.
CHAPTER THREE METHODOLOGY
3.0 Introduction
The effect of the leech therapy on body composition is a basic research on determining the difference in impedance measurement before and after the treatment.
The process of the study can be depicted by the flowchart in Figure 3.1 below.
Figure 3.1: Flowchart of study
Preparation for demographic data and measurement setup at leech therapy treatment center in Pantai Dalam, KL
Completion of data by subjects with 2 forms:
i- Personal information ii- Consent form
Conducting measurement for pre-treatment data
Keying in information and data measurement before leech therapy
Completion of treatment within 1 hour
Keying in information and measurement after treatment of leech therapy
Analyzing data using the statistical tool
32 3.1 Subjects / patient
The criteria of the subjects of this study are constrained by the method of the analysis which uses the BIA measurement and the treatment of leech therapy. For BIA measurement, a subject should not be in the early pregnancy stage or implanted with any electronic devices such as a pace maker. Moreover, leech therapy treatment also could not be performed on pregnant women due to the influence of treatment in the course of developing pregnancy and sometimes causing miscarriage. In addition, the leech therapy patient must not experiencing any symptoms of low blood pressure, anemia or anything similar as well as taking any medication that has the same effect as Hirudin enzyme. This will make the conditions of the patient become worse and thus is not eligible to be the subject of this study.
Overall, there are 50 patients that agree to take part in this study. All information regarding this study was brief thoroughly.
3.2 Measurement setup
Before measurement begins, the patient must be sure to be in the supine position.
The measurement is taken from the right hand and right foot of the patient. So, the placement of the electrode on proximal part is placed on the dorsal surface of the wrist so that the upper border of the electrode bisects the styloids processes of the ulna and radius and dorsal surface of the ankle that it able to bisects medial and lateral malleoli.
In addition, the right foot and right hand where the distal placement of electrodes is done at the base of the second or third metacarpal phalangeal joints. These distance at the proximal and distal are always check to be in the distance of at least 5cm.
Then, the clip wire is attached to the self adhesive disposable electrodes to take the measurement. Basically, there are 2 main wires that are used which are red and
black. The position of the red wire is placed at the distal and black is always positions at the proximal distance. This can be illustrated below.
Figure 3.2 Placement of electrodes at hand
Figure 3.3 Placement of electrodes at leg
3.3 Data measurement
The procedure could be outline as below:
1. After the patient is in the correct position, then the body must be check so that it won’t touch one and other. This is to avoid any short circuit in the path of the current inside the body
2. The electrodes is placed on the correct position
3. The crocodile clips is attached to the metal tab strip of the electrode.
4. The bioimpedance machine which is Quadscan 4000 is switch on.
34 5. All the data from the patient were insert one by one before the measurement start.
6. After 3 to 4 minutes, the measurement start to take place and results are display on the LCD screen.
7. Then the data could be downloaded via Bluetooth connection at any time for analysis.
3.4 Statistical analysis
This analysis is divided into a few parts that are bar graphs, descriptive statistics, independent t-test, paired sample t-test and ANOVA. The steps are depicted more on appendix A.
3.4.1 Bar graph
The step by step procedures are by clicking this one by one.
Analyze descriptive statistics frequencies chart.
Since the bar graph is set in percentage, all the frequencies given are automatically converted.
3.4.2 Descriptive statistics This can be done as follows.
Analyze descriptive statistics frequencies descriptives options ok.
All the important descriptive parameters are chosen for the analysis. These include mean, standard mean error, kurtosis, variance and standard deviation.
3.4.3 Independent sample t-test
Following the order below, the results are obtained from independent t-test.
Analyze compare means independent sample t-test ok The entire test variables are put on the box and grouping variable is defined.
3.4.4 Paired sample t-test
Following the order below, the results are obtained from paired sample t-test.
Analyze compare means paired sample t-test ok The entire paired variables are put on the box for analysis.
3.4.5 ANOVA
This can be done as follows.
Analyze compare means one way ANOVA post hoc ok
36 CHAPTER FOUR
RESULTS
4.0 Statistical analysis
The data that were collected from the Quadscan 4000 are all analyzed closely by using the SPSS tool. The involve test are pair t-test, independent sample t-test and ANOVA which stands for analysis of variance. The effect of body composition is measured based in the bioimpedance parameter which are divided into 2 called as primary and secondary bioimpedance parameter. The parameter involves were listed in Table 4.0 as below.
Table 4.0: Bioimpedance parameter
Main bioimpedance parameter Secondary bioimpedance parameter
Impedance at 5K Percentage of fat before test
Impedance at 50K Min fat before test
Impedance at 100K Max fat before test
Impedance at 200K Percentage of lean before test
Reactance at 50K Min lean before test
Resistance at 50K Max lean before test
Phase Angle at 50 kHz Dry lean weight before test Basal metabolic rate Percentage of water before test
Intracellular water Min water before test
Normal value of intra cellular water Max water before test Extra-cellular water Density before test Normal value of extra cellular water Nutrition before test
Total body water volume Normal nutrition before test Min total body water Third space value before test Max total body water volume before test
Body cell mass before test
4.1 Demographic data
The demographic data is basically a requirement for the patient to complete in order to enrol in this study. Important background information were listed such as gender, marital status, occupation, age, illness, number of treatment and subject’s daily activity. BMI measurements are derived from the demographic information and compare with the analysis data from Quadscan 4000.
Figure 4.1: Bar graph for subject’s gender
Based on figure 4.1, gender of patient can be classified for the leech therapy treatment that is dominated by female by 54% of the participant of this study which is more than half. Therefore, the male participants were identified to be only 46% from the 50 patients that enrolled. These participants were randomly picked from the patients who come to get the leech therapy treatment in the treatment center. Thus, this shows that this treatment is more popular among females than males by 8% difference.
38 Figure 4.2: Bar graph for subject’s marital status
For the above figure 4.2, the variables involve is the marital status of the subjects. The bar graph shows that the patients that come to get the treatment are mostly single person rather than married. This is depicted by the value of single patient that covers 60% than the total patient that come to get treatment. Married person that are chosen were only 20 persons that brings the statistical percentage up to 40% only.
Figure 4.3: Bar graph for range of subject’s age
From figure 4.3, the ranges of subject’s age are determined from the maximum and minimum value of the data. Using cross tabulation method, there are 5 significance ranges which dominated by the patient from age 21-30 years by 36%. This is followed by those who are younger in the category of age less than 21 by 24%. Then, the percentage is decreases from age of more than 50 by 18%, 41 to 50 by 12% and least participant for range of 31 to 40 years by only 10%.
40 Figure 4.4: Bar graph for subject’s occupation
From the chart of figure 4.4, most of the subjects are working in the private sector showing 42% participation rate where those who fall in this category are consultant, driver, secretary, artist, lecturer, executive, technician, manager, banker, clerk, engineer and hotelier. However, 38% out of 50 subjects were categorized as students where this covers student from primary to tertiary level of education. The public sector is fall behind with 6% in which subjects currently working as nurse or research assistant in the public university. Furthermore, unemployed participants were categorized as housewife or retiree with overall 12% from the total patients. Self- working subject is only one where it contributes 2% of the total statistics.
Figure 4.5: Bar graph for number of leech therapy treatment
The bar graph is indicating that most of the participants were a first timer in getting a leech therapy treatment with 88% and the rest were the second time getting this treatment and third time with percentages 10% and 2% respectively.
42 Figure 4.6: Bar graph for subject’s daily activity
The above figure indicating that majority of the subject is regarded as having a medium level of daily activity with 64% from total statistics. However, those who fall under low to medium and medium to high level of activity were identified as 10% and 20% respectively. This is trailed by another two category that are activity level is considered very low with 2 % and double by 4% for very high level of activity.
Figure 4.7: Bar graph for BMI readings
The BMI readings were taken before and after the leech therapy treatment and most of the participant of this study are having a BMI from 20 to 25 with 46% in total.
The BMI range from 26 to 30 was more with 20% than BMI range of less than 20 that is only 18%. About 6% of total patient is having BMI of more than 35.
44
46
48
50
52
54
56
58
60
62
64
66
68
70
72
74
4.4 Comparison of bioimpedance parameter
This is the analysis to see the comparison of bioimpedance parameter with the demographic variable namely age and BMI groups
4.4.1 Comparison of Bioimpedance Parameters with Patient’s Age Group.
In this section Comparison of Bioimpedance Parameters with Patient’s Age Group are shown here. For ease of reporting and analysis age group also divided into four groups here. For analysis the bioimpedance parameters are divided into two groups here. They are primary and secondary bioimpedance parameters. Also need to consider the condition before and after treatment here. Independent T test and ANOVA test was used here for this analysis. The value of P should be less than 5% for significance.
The age group 2 until age group 5 have almost the same pattern in age group 1 and some have some missing value because insufficiency of parameters for analysis.
Table 4.30 Comparisons of Main Bioimpedance Parameters before Treatment for Female age group 1.
Main Bioimpedance Parameters Overall (p) BMI (P) Diseases Grp(p) Impedance value at 5KHz 0.000 0.000 0.000
Impedance value at 50KHz 0.000 0.000 0.000 Impedance value at 100KHz 0.000 0.000 0.000 Impedance value at 200KHz 0.000 0.000 0.000
Reactance at 50KHz 0.002 0.000 0.000
Resistance at 50KHz 0.000 0.000 0.000 Phase Angle at 50 KHz 0.000 0.000 0.000 Basal metabolic rate 0.099 0.000 0.000 Intracellular water 0.720 0.000 0.000 Normal value of intracellular
water
0.345 0.000 0.000 Extra-cellular water 0.534 0.000 0.000 Normal extra cellular water 0.710 0.000 0.000 Total body water volume 0.749 0.000 0.000 Min total body water volume 0.011 0.000 0.000 Max total body water volume 0.182 0.000 0.000
Body cell mass 0.724 0.000 0.000
* No of treatment does not have enough parameter for analysis.
76 Table 4.31 Comparisons of Secondary Bioimpedance Parameters before
Treatment for Female Age group 1.
Secondary Bioimpedance Parameters Overall (p) BMI (p)
Diseases Grp(p)
Percentage of fat 0.294 0.000 0.000
Min fat 0.186 0.000 0.001
Max fat 0.439 0.000 0.001
Percentage of lean 0.294 0.000 0.000
Min lean 0.244 0.000 0.000
Max lean 0.246 0.000 0.000
Dry lean weight 0.049 0.000 0.004
Percentage of water 0.575 0.000 0.000
Min water 0.011 0.000 0.000
Max water 0.182 0.000 0.000
Density 0.852 0.000 0.000
Nutrition 0.001 0.000 0.000
Normal nutrition - 1.000 1.000
Third space 0.000 0.000 0.000
* No of treatment does not have enough parameter for analysis.
Table 4.32 Comparisons of Main Bioimpedance Parameters after Treatment for Female Age group 1.
Main Bioimpedance Parameters Overall (p) BMI (P) Diseases Grp(p) Impedance value at 5KHz 0.001 0.000 0.000
Impedance value at 50KHz 0.001 0.000 0.000 Impedance value at 100KHz 0.002 0.000 0.000 Impedance value at 200KHz 0.003 0.000 0.000
Reactance at 50KHz 0.007 0.000 0.000
Resistance at 50KHz 0.002 0.000 0.000 Phase Angle at 50 KHz 0.058 0.000 0.000 Basal metabolic rate 0.079 0.000 0.000 Intracellular water 0.693 0.000 0.000 Normal value of intracellular
water
0.345 0.000 0.000 Extra-cellular water 0.753 0.000 0.000 Normal extra cellular water 0.710 0.000 0.000 Total body water volume 0.582 0.000 0.000 Min total body water volume 0.011 0.000 0.000 Max total body water volume 0.182 0.000 0.000
Body cell mass 0.698 0.000 0.000
* No of treatment does not have enough parameter for analysis.
Table 4.33 Comparisons of Secondary Bioimpedance Parameters after Treatment For Female Age group 1.
Secondary Bioimpedance Parameters Overall (p) BMI (p)
Diseases Grp(p)
Percentage of fat 0.246 0.000 0.000
Min fat 0.235 0.000 0.005
Max fat 0.633 0.000 0.000
Percentage of lean 0.246 0.000 0.000
Min lean 0.160 0.000 0.000
Max lean 0.204 0.000 0.000
Dry lean weight 0.047 0.000 0.005
Percentage of water 0.347 0.000 0.000
Min water 0.011 0.000 0.000
Max water 0.182 0.000 0.000
Density 0.852 0.000 0.000
Nutrition 0.000 0.000 0.000
Normal nutrition - 1.000 1.000
Third space 0.001 0.000 0.000
* No of treatment does not have enough parameter for analysis.
Table 4.34 Comparisons of Main Bioimpedance Parameters before Treatment for male age group 1.
Main Bioimpedance Parameters Overall (p) BMI (P) No of
treatment (p)
Diseases Grp(p) Impedance value at 5KHz 0.000 0.104 0.640 0.000 Impedance value at 50KHz 0.000 0.024 0.688 0.001 Impedance value at 100KHz 0.000 0.022 0.640 0.001 Impedance value at 200KHz 0.000 0.024 0.574 0.000
Reactance at 50KHz 0.002 0.001 0.207 0.154
Resistance at 50KHz 0.000 0.023 0.686 0.001
Phase Angle at 50 KHz 0.000 0.003 0.280 0.001
Basal metabolic rate 0.099 0.000 0.194 0.322
Intracellular water 0.720 0.000 0.324 0.142
Normal value of intracellular water
0.345 0.000 0.217 0.781
Extra-cellular water 0.534 0.000 0.211 0.089
Normal extra cellular water 0.710 0.000 0.217 0.790 Total body water volume 0.749 0.000 0.246 0.057 Min total body water volume 0.011 0.000 0.232 0.769 Max total body water volume 0.182 0.000 0.209 0.799
Body cell mass 0.724 0.000 0.328 0.140
